(1) A method for determination of the free mobility (surface net charge) of macromolecules based on the Ferguson plot in agarose-polyacrylamide copolymers was developed. The method, for the first time, avoids the error due to the assumption of Ferguson plot linearity in the lowest gel concentration range (O to 3.5% total acrylamide (T)). It thereby recaptures the key advantage of gel electrophoresis over chromatography in the physical characterization of macromolecules, i.e., the simultaneous evaluation of surface net charge and size of macromolecules. (2) A discontinuous (moving boundary electrophoresis) buffer system for DNA fragments in the size range of 8 to 23,000 bp was developed which by replacing the characteristic absolute mobilities by relative ones is capable of reducing the error in zone identification. (3) A method for polyacrylamide gel electrophoresis of DNA fragments, 0.3 to 23 kbp in size, using gel 15% crosslinked with DATD was developed which yields superior resolution compared to agarose gels conventionally used in that size range. (4) The dependence of the electrophoretic mobility of DNA fragments on ethidium bromide concentration used in prelabeling, on temperature and ionic strength was determined. (5) A method for evaluating polydisperse gel patterns yielding size, net charge and relative concentration at each point in the pattern was developed. The method advances one previously developed in application to semisynthetic protein conjugates with bacterial coat carbohydrates of importance in vaccine production, by being able to assign relative concentrations to the components characterized by size and charge. (6) A gel electrophoresis apparatus for thin-layer, horizontal gel strips submerged under inert solvents while applicable to discontinuous buffer systems was constructed. The apparatus is the first to combine "stacking' of large sample volumes, "submarine" operation which prevents gel dehydration during electrophoresis, and reduction of load from the microgram to the nanogram level. (7) A technique of electrofocusing in immobilized pH gradients, using a gel submerged under silicone oil and thermostated by Peltier cells was developed. The method prevents water exudates on the gel surface and dehydration without loss of efficient temperature control, and therefore allows for prolonged electrofocusing frequently required for attaining the isoelectric endpoint of a protein.